Synthetic pyrethroids, an important class of neurotoxic insecticides used worldwide in agriculture and public health, cause toxic effects in animals by modifying the normal function of voltage-sensitive sodium channels. Sodium channels in mammalian tissues exist as multiple isoforms that exhibit differential tissue distribution and developmental regulation and are encoded by members of a multigene family. Previous studies of pyrethroid action have not considered the differential sensitivity of sodium channel isoforms as a determining factor in the toxicity of these compounds. The proposed research will test the hypothesis that differences in the pyrethroid sensitivity of mammalian sodium channel isoforms are important determinants of the nature and severity of pyrethroid intoxication. Specific experimental aims are: (1) to define the actions of pyrethroid insecticides on cloned rat sodium channel isoforms expressed in frog (Xenopus laevis) oocytes or transfected mammalian cells;(2) to compare the actions of pyrethroids on orthologous rat and human sodium channel isoforms expressed in Xenopus oocytes or trnasfected mammalian cells;and (3) to map the the structural determinants of pyrethroid action by the construction, functional expression and pharmacological characterization of specifically mutated sodium channels. Cloned sodium channel isoforms will be expressed in Xenopus oocytes or transiently transfected mammalian cells (e.g., HEK-293) and the actions of pyrethroid insecticides on expressed channels will be assessed using electrophysiological recordings of macroscopic sodium currents under two-electrode voltage-clamp or whole cell patch-clamp conditions. Structural determinants of pyrethroid sensitivity will be identified by site-directed mutagenesis, expression, and pharmacological comparison of native a mutated sodium channel isoforms. Results of these studies will define the role of different sodium channel isoforms in pyrethroid toxicity, establish whether rat sodium channels are valid toxicological models for the sensitivity of human sodium channels to pyrethroids, and identify sodium channel domains that determine pyrethroid sensitivity.
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